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P J Mitchell

Publications and source records attributed to P J Mitchell.

At least 73 records · Page 4Linked to original sources
PubMed1995

Expression patterns of the novel receptor-like tyrosine kinase, DDR, in human breast tumours.

Using a reverse transcriptase-polymerase chain reaction based differential screening procedure, we have identified the discoidin domain receptor as a protein tyrosine kinase that is expressed in lymph nodes containing breast tumour metastases. By Northern blotting and in situ hybridisation we have demonstrated the expression of the discoidin domain receptor in human primary breast tumour samples, metastasis-containing lymph nodes and a number of normal tissues. Direct comparison of malignant breast and adjacent normal epithelial tissue revealed over expression in the tumour cells.

Amino Acid Sequence↗
PubMed1995

Alternative mRNAs encode multiple isoforms of transcription factor AP-2 during murine embryogenesis.

Transcription factor AP-2 has been implicated as an important regulator of gene expression during vertebrate embryogenesis. We report here the cDNA cloning and analysis of mouse embryonic mRNA splice variants encoding four AP-2 isoforms. Isoform 1 is the homolog of the previously known human (HeLa) AP-2. The three new AP-2 isoforms all share the same DNA binding/dimerization domain as isoform 1 but either lack the proline-rich transcriptional activation domain encoded by exon 2 (isoform 2) or have different amino-termini encoded by two previously unknown alternative first coding exons for AP-2 (isoforms 3 and 4). All four AP-2 mRNA variants are present at significant levels between Days 11.5 and 17.5 of mouse embryogenesis. Variants 1, 3, and 4 show qualitatively but not quantitatively similar restricted expression patterns in 8.5-12.5 dpc embryos examined by in situ hybridization. At mid-embryogenesis, variant 3 is the major AP-2 mRNA species in the nervous system and in total embryo RNA but is less prevalent than variants 1 and 4 in the epidermis. The four mRNAs are all induced, although unequally, during differentiation of P19 cells into neural cell types and by cAMP stimulation of primary astrocytes. Variants 1-3 are coexpressed in different ratios in HeLa cells and in three human glioblastoma cell lines. These findings reveal that transcriptional regulation by AP-2 is likely to be more complex than previously assumed given the potential for multiple AP-2 homo- and heterodimeric DNA binding forms.

Alternative Splicing↗
PubMed1995

The therapeutic efficacy and prophylactic activity of doramectin against Dictyocaulus viviparus in cattle.

Three groups of 8, 4-month-old male Jersey or Jersey-cross calves were infected with 2400 Dictyocaulus viviparus L3 larvae and either left untreated or injected subcutaneously with 200 micrograms/kg doramectin 5 or 25 days after infection (DAI). Lungworms were found in all untreated cattle (geometric mean = 49) at necropsy 39 or 40 DAI. None was found in any of the treated cattle. In a second experiment, groups of 6, 8-month-old calves were untreated or injected with 200 micrograms/kg doramectin 28, 21 or 14 days before each calf was challenged with 2700 D viviparus larvae. Lungworms were recovered at necropsy 32 to 34 DAI. The geometric mean worm burden in the untreated cattle was 550. This was reduced by 100%, 99.5% and 94.1% in calves treated with doramectin 14, 21 or 28 days, respectively, before infection. It was concluded that doramectin is a highly effective anthelmintic against D viviparus adult or L4 infections of cattle, and that reinfection of treated cattle will be significantly reduced for at least 28 days after treatment.

Animals↗
PubMed1995

Human cytomegalovirus (HCMV) immediate-early enhancer/promoter specificity during embryogenesis defines target tissues of congenital HCMV infection.

Congenital human cytomegalovirus (HCMV) infection is a common cause of deafness and neurological disabilities. Many aspects of this prenatal infection, including which cell types are infected and how infection proceeds, are poorly understood. Transcription of HCMV immediate-early (IE) genes is required for expression of all other HCMV genes and is dependent on host cell transcription factors. Cell type-specific differences in levels of IE transcription are believed to underlie differences in infection permissivity. However, DNA transfection experiments have paradoxically suggested that the HCMV major IE enhancer/promoter is a broadly active transcriptional element with little cell type specificity. In contrast, we show here that expression of a lacZ gene driven by the HCMV major IE enhancer/promoter -524 to +13 segment is restricted in transgenic mouse embryos to sites that correlate with known sites of congenital HCMV infection in human fetuses. This finding suggests that the IE enhancer/promoter is a major determinant of HCMV infection sites in humans and that transcription factors responsible for its regulation are cell type-specifically conserved between humans and mice. The lacZ expression patterns of these transgenic embryos yield insight into congenital HCMV pathogenesis by providing a spatiotemporal map of the sets of vascular, neural, and epithelial cells that are likely targets of infection. These transgenic mice may constitute a useful model system for investigating IE enhancer/promoter regulation in vivo and for identifying factors that modulate active and latent HCMV infections in humans.

Animals↗
PubMed1995

Crossed cerebellar diaschisis and brain recovery after stroke.

BACKGROUND AND PURPOSE: Although crossed cerebellar diaschisis is well recognized after stroke, there is controversy concerning its clinical correlations and serial changes, and little is known about its prognostic value. METHODS: We studied crossed cerebellar diaschisis and cerebral hypoperfusion in 47 patients with acute middle cerebral cortical infarction using 99mTc-hexamethylpropyleneamine oxime and single-photon emission computed tomography within 72 hours of stroke onset. Thirty-one of these patients had outcome studies at 3 months; 15 of the 31 underwent an additional scan after acetazolamide injection. Tissue loss was determined with computed tomography, performed at outcome in 28 patients. Clinical stroke severity was assessed with the Canadian Neurological Scale and Barthel Index. Cerebellar blood flow asymmetry was studied in 22 healthy, age-matched control subjects. RESULTS: Cerebellar blood flow asymmetry was significant in patients (mean +/- SE, 9.76 +/- 0.78%; P < .001) but not in control subjects (-0.22 +/- 0.56%). Crossed cerebellar diaschisis was strongly associated with infarct hypoperfusion volume at both acute (regression coefficient +/- SEb, b = 6.76 +/- 0.65; P < .001) and outcome stages (b = 6.13 +/- 0.63; P < .001). Cross-sectionally over the first 72 hours, infarct hypoperfusion volume decreased by 2% for each hour from onset (P < .05), while crossed cerebellar diaschisis remained unchanged. Canadian Neurological Scale score at the acute stage was negatively associated with acute crossed cerebellar diaschisis (b = -0.10 +/- 0.05; P < .05) after allowing for infarct hypoperfusion volume. Crossed cerebellar diaschisis did not change between acute-stage, outcome, and postacetazolamide scans. Acute-stage crossed cerebellar diaschisis predicted outcome Barthel Index score (b = -0.28 +/- 0.14; P = .05) and tissue loss (b = 3.81 +/- 0.96; P < .001) but was no longer an independent prognostic factor after allowing for acute-stage infarct hypoperfusion volume. CONCLUSIONS: This study shows that crossed cerebellar diaschisis is a functional phenomenon that correlates with both stroke severity and infarct hypoperfusion volume and persists despite neurological recovery. Although acute-stage crossed cerebellar diaschisis has no prognostic value independent of acute-stage hypoperfusion volume, it might indicate the proportion of nutritional to nonnutritional perfusion at the infarct site and hence be useful in the evaluation of reperfusion therapies in the acute stage.

Acetazolamide↗
PubMed1995

Cerebral venography and manometry in idiopathic intracranial hypertension.

Cerebral venography and manometry in nine patients with idiopathic intracranial hypertension consistently showed venous hypertension in the superior sagittal sinus and proximal transverse sinuses, with a significant drop in venous pressure at the level of the lateral third of the transverse sinus. The abnormality, clearly demonstrated by manometry, was not well shown on the venous phase of cerebral angiography. The appearance of the transverse sinus on venography varied from smooth tapered narrowing to discrete intraluminal filling defects that resembled mural thrombi. Two patients with intracranial hypertension due to minocycline did not show venous hypertension.

Adolescent↗
PubMed1995

Detection of discrete white matter lesions after irreversible compression of MR images.

PURPOSE: To validate the use of techniques of irreversible compression of images, which can be performed using a block-based discrete cosine transform technique as defined by the Joint Photographic Experts Group, before they can be used in clinical applications, by evaluating the effect of compression on the ability of observers to detect discrete white matter lesions on MR images of the brain. METHODS: Sixty T2- and intermediate-weighted spin-echo images were compressed with varying degrees of coefficient quantization with compression ratios from 1:1 to more than 40:1, randomized, and evaluated by three observers blinded to the degree of compression. RESULTS: No significant difference in the number of lesions detected was apparent until compression ratios reached 40:1, despite a significant subjective loss in perceived image quality at 20:1. Only small (< or = 5 mm) lesions were missed at the highest degree of compression. No significant differences were observed in the detection of confluent periventricular white matter disease at any degree of compression tested. CONCLUSIONS: The use of high degrees of irreversible compression of MR images may be acceptable for diagnostic tasks.

Brain↗
PubMed1994

Cell type-specific regulation of expression of transcription factor AP-2 in neuroectodermal cells.

AP-2 is a cell type-specific DNA-binding transcription factor that regulates selected target genes in vertebrate organisms. Here we investigated cell type-specific expression and regulation of AP-2 in neuroectodermal cell lineages. During retinoic acid (RA)-mediated differentiation of P19 embryonal carcinoma cells into neuroectodermal cell types that include immunohistochemically defined neurons and astrocytes, we observed a strong induction of AP-2 transcripts and protein. In contrast, AP-2 mRNA was not induced in P19 cells which undergo mesoendodermal differentiation in response to 1% dimethylsulfoxide or low concentrations of RA, respectively. The potential of both neurons and astrocytes to express AP-2 was ascertained by using cerebellar neurons and astrocytes derived from newborn mice. Unlike these types of cells, microglial cells do not express AP-2. Dibutyryl cyclic AMP further enhanced levels of AP-2 transcripts in both P19 astrocytes and primary astrocytes which also respond to agents elevating intracellular cAMP (noradrenaline, isoproterenol, forskolin). The cAMP-dependent induction of AP-2 could be blocked by inhibitors of protein kinase A. In contrast to its action in P19 cells, RA had no effect on AP-2 mRNA levels in primary astrocytes. Our results indicate that AP-2 may play a role as a retinoic acid-sensitive regulator during differentiation of neurons and glia from an embryonic neural precursor. Furthermore, AP-2 may be involved in gene transcription in both mature neurons and astrocytes.

Animals↗
PubMed1994

Tpr, a large coiled coil protein whose amino terminus is involved in activation of oncogenic kinases, is localized to the cytoplasmic surface of the nuclear pore complex.

From a panel of monoclonal antibodies raised against fractions of rat liver nuclear envelopes (NEs), we have identified an antibody, RL30, which reacts with novel nuclear pore complex (NPC) antigens that are not O-glycosylated. By immunofluorescence staining of cultured cells, RL30 reacts exclusively with the NE in a punctate pattern that largely coincides with that of identified NPC proteins. RL30 labels only the cytoplasmic surface of the NPC in immunogold electron microscopy, predominantly in peripheral regions nearby the cytoplasmic ring. In immunoblots of isolated rat liver NEs and cultured rat cells, RL30 recognizes a 265-kD band, as well as a series of 175-265-kD bands in rat liver NEs that are likely to be proteolytic products of p265. Sequencing of peptides from the 175- and 265-kD RL30 antigens of rat liver revealed that they are both closely related to human Tpr, a protein whose amino-terminal 150-250 amino acids appear in oncogenic fusions with the kinase domains of the met, trk, and raf protooncogenes. We found that in vitro translation of human Tpr mRNA yields a major 265-kD band. Considered together, these data indicate that the 265-kD RL30 antigen in the NPC is the rat homologue of Tpr. Interestingly, Tpr contains an exceptionally long predicted coiled coil domain (approximately 1600 amino acids). The localization and predicted structure of Tpr suggest that it is a component of the cytoplasmic fibrils of the NPC implicated in nuclear protein import. Immunofluorescence microscopy shows that during NPC reassembly at the end of mitosis, Tpr becomes concentrated at the NE significantly later than O-linked glycoproteins, including p62. This indicates that reassembly of the NPC after mitosis is a stepwise process, and that the Tpr-containing peripheral structures are assembled later than p62.

Amino Acid Sequence↗
PubMed1994

Cloning and characterisation of cDNAs encoding a novel non-receptor tyrosine kinase, brk, expressed in human breast tumours.

Using a polymerase chain reaction based differential screening approach, we have isolated and characterised a cDNA from a human metastatic breast tumour representing a novel protein tyrosine kinase (brk). Sequencing of brk cDNAs isolated from T-47D and MCF-7 human breast tumour cell lines indicate that they encode a protein with the features of a novel nonreceptor tyrosine kinase, including amino terminal SH3 and SH2 domains. When synthesised in recombinant baculovirus and bacterial expression systems, brk protein products are capable of autophosphorylation on tyrosine residues. Initial expression studies have detected low levels of brk transcripts in some human breast tumours and breast tumour cell lines, but not in normal breast tissue.

Amino Acid Sequence↗
PubMed1993

Use of PC12 cells as a neurotoxicological screen: characterization of anticyanide compounds.

A series of six biochemical markers of cyanide toxicity (dopamine release, hydroperoxide generation, cytosolic-free calcium levels, catalase activity, cytochrome oxidase activity, and superoxide dismutase activity) in cultured rat pheochromocytoma (PC12) cells were used to establish a screen for evaluation of potential anticyanide compounds. Thirty-nine substances, including anticonvulsants, adrenergic blockers, antioxidants, and antipsychotics were tested and ranked according to the results. Based on the composite scoring in all six assays, carbamazepine, mannitol, allopurinol, and phenytoin were ranked as the most effective anticyanide compounds. Additionally, known cyanide antidotes (e.g., pyruvate, mercaptopyruvate, alpha-ketoglutarate, naloxone, and flunarizine) obtained relatively high ranking in the PC12 cell screen. Furthermore, a significant correlation was found between protective effects (based on LD50s) of cyanide antidotes in mice and ranking in the in vitro screen. This study illustrates that by assaying a series of biochemical markers in a neuronal-type cell line, a rapid, cost-effective in vitro toxicological screen is possible. Several compounds have been identified which inhibit the biochemical effects of cyanide and may be used to enhance effectiveness of the standard cyanide antidotes.

Animals↗
PubMed1993

Venlafaxine exhibits pre-clinical antidepressant activity in the resident-intruder social interaction paradigm.

Venlafaxine, a novel 2-phenyl-2-(1-hydroxycycloalkyl) ethylamine, is a potent inhibitor of 5-hydroxytryptamine and noradrenaline reuptake and exhibits a profile of activity in pre-clinical in vitro biochemical studies predictive of antidepressant activity. The studies described here examined the effects of acute and chronic treatment with venlafaxine on the behaviour of resident rats confronted with an unfamiliar, non-treated, intruder conspecific. Ethological analysis of the social encounters revealed that acute, subcutaneous, treatment with venlafaxine, 20-180 mumol kg-1, induced a selective, dose-related, reduction in aggressive behaviour (ID50 = 24.87 mumol kg-1) concomitant with increased flight behaviour. In contrast, chronic treatment with venlafaxine, 20 mumol kg-1 day-1, via subcutaneously-implanted osmotic mini-pumps, induced a marked elevation in aggressive behaviour concomitant with reduced flight behaviour. These diametrically opposite effects of acute and chronic venlafaxine treatment on the agonistic behaviour of resident rats are consistent with the behavioural effects of similar treatment regimes previously identified for a range of antidepressant drugs that differ widely in their acute pharmacology. These data strongly support the potential antidepressant activity of venlafaxine and are consistent with the results of recent clinical trials which demonstrate that venlafaxine exhibits significant antidepressant activity.

Aggression↗
PubMed1992

Effects of caffeine, time of day and user history on study-related performance.

The individual and interactive effects of caffeine, time of day and history of caffeine consumption on several study-related tasks were investigated in 25 subjects (6 males, 19 females). Performance was measured on short term memory (STM), mental arithmetic (MA), reading comprehension, serial search (SS) and verbal reasoning (VR). Subjects attended eight experimental sessions, at four times of day (0100, 0700, 1300 and 1900 hours), after ingesting caffeine (4 mg/kg) or placebo. Subjects were assigned to a low, moderate or high user group on the basis of a caffeine consumption questionnaire. Reading comprehension was affected by time of day, while caffeine improved performance on all mental speed-related tasks. High caffeine users performed more poorly than other groups on the verbal reasoning task. Several interactions between the three independent variables were observed on a number of tasks, supporting the contention that different processes underlying various types of cognitive performance are differentially, and often jointly, affected by caffeine, time of day and user history. Implications of caffeine usage on academic performance were discussed.

Adult↗
PubMed1992

The human tpr gene encodes a protein of 2094 amino acids that has extensive coiled-coil regions and an acidic C-terminal domain.

We recently provided evidence that the human tpr gene encodes a 726 amino acid protein (designated tpr-S) and identified an alternatively spliced transcript that encodes a larger tpr protein with an extended C-terminal domain. In this study, through isolation and sequencing of tpr cDNA clones, we have established that this alternatively spliced transcript encodes a protein of 2094 amino acids (designated tpr-L). The larger tpr protein is predicted to have extensive regions of alpha-helix and several stretches of a heptad repeat that is characteristic of proteins adopting a coiled-coil conformation. Furthermore the carboxy domain of this protein is very rich in acidic residues and exhibits homology (58-80%) to the acidic regions of several nuclear proteins, including the Drosophila engrailed protein, Escherichia coli RNA polymerase sigma subunit and nucleolin. To gain additional insight into the function of tpr we examined the expression of tpr transcripts in tissues from adult rat. The highest levels of tpr transcripts were observed in testis, lung, thymus and spleen.

Amino Acid Sequence↗
PubMed1992

Nucleotide sequence analysis of human tpr cDNA clones.

In this study we have characterized cDNA clones corresponding to a gene, called tpr, that has been implicated in the activation of the met and raf proto-oncogenes. Sequencing of tpr clones isolated from an HT1080 human fibrosarcoma cell line cDNA library identified an open reading frame (ORF) of 726 amino acids. In addition we have established that alternative splicing can result in the deletion of a 30 bp sequence that spans the translation termination site of this ORF. This modification generates mRNAs encoding a tpr protein that has an extended C-terminal domain. The 726 amino acid tpr protein is predicted to have extensive regions of alpha-helix and has three stretches of a heptad repeat motif that is characteristic of proteins adopting a coiled-coil conformation. The tpr protein exhibits weak homology (28-39%) to the alpha-helical domains of several proteins including tropomyosin, spectrin, laminin B1, the Drosophila glued protein and the tail region of myosin heavy chain.

Amino Acid Sequence↗
PubMed1992

Myasthenia gravis in pregnancy.

OBJECTIVE: Our purpose was to review our experience with nine cases of myasthenia gravis complicating pregnancy. METHODS: We reviewed the records of all patients discharged from the Vancouver Salvation Army Grace Hospital from January 1979 to January 1989 with an admission diagnosis of myasthenia gravis in pregnancy. RESULTS: Nine patients delivered 11 infants. Four suffered antepartum exacerbations. The most severe of these, respiratory failure, occurred in a thymectomized patient. Labor was rapid and uncomplicated except for a single case of cephalopelvic disproportion delivered abdominally. We noted no puerperal exacerbations. Our solitary case of neonatal myasthenia gravis was managed in the special care nursery with intramuscular neostigmine given over a 16-day period. CONCLUSION: There are no predictive factors that identify the mother at risk for peripartum exacerbation of myasthenia gravis or the infant at risk for neonatal myasthenia gravis. Optimal care for the parturient is achieved through a team approach involving the obstetrician, neurologist, and pediatrician.

Adult↗